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R Bouillon

Publications and source records attributed to R Bouillon.

At least 253 records · Page 14Linked to original sources
PubMed1991

Photoaffinity labeling of human serum vitamin D binding protein and chemical cleavages of the labeled protein: identification of an 11.5-kDa peptide containing the putative 25-hydroxyvitamin D3 binding site.

In this paper, we describe photoaffinity labeling and related studies of human serum vitamin D binding protein (hDBP) with 25-hydroxyvitamin D3 3 beta-3'-[N-(4-azido-2-nitrophenyl)amino]propyl ether (25-ANE) and its radiolabeled counterpart, i.e., 25-hydroxyvitamin D3 3 beta-3'-[N-(4-azido-2-nitro-[3,5-3H]phenyl)amino]propyl ether (3H-25-ANE) (Ray et al., 1986, 1991). We have carried out studies to demonstrate that (1) 25-ANE competes with 25-OH-D3 for the binding site of the latter in hDBP and (2) 3H-25-ANE is capable of covalently labeling the hDBP molecule when exposed to UV light. Treatment of a sample of purified hDBP, labeled with 3H-25-ANE, with BNPS-skatole produced two Coomassie Blue stained peptide fragments, and the majority of the radioactivity was associated with the smaller of the two peptide fragments (16.5 kDa). On the other hand, cleavage of the labeled protein with cyanogen bromide produced a peptide (11.5 kDa) containing most of the covalently attached radioactivity. Considering the primary amino acid structure of hDBP, this peptide fragment (11.5 kDa) represents the N-terminus through residue 108 of the intact protein. Thus, our results tentatively identify this segment of the protein containing the binding pocket for 25-OH-D3.

Affinity Labels↗
PubMed1991

Synthesis of 25-hydroxyvitamin D3 3 beta-3'-[N-(4-azido-2-nitrophenyl)amino]propyl ether, a second-generation photoaffinity analogue of 25-hydroxyvitamin D3: photoaffinity labeling of rat serum vitamin D binding protein.

Vulnerability of 25-hydroxy-[26,27-3H]vitamin D3 3 beta-N-(4-azido-2-nitrophenyl)glycinate, a photoaffinity analogue of 25-hydroxyvitamin D3 (25-OH-D3) (Ray et al., 1986) toward standard conditions of carboxymethylation promoted us to synthesize 25-hydroxyvitamin D3 3 beta-3'-[N-(4-azido-2-nitrophenyl)amino]propyl ether (25-ANE), a hydrolytically stable photoaffinity analogue of 25-OH-D3, and 25-hydroxyvitamin D3 3 beta-3'-[N-(4-azido-2-nitro-[3,5-3H]phenyl)amino] propyl ether (3H-25-ANE), the radiolabeled counterpart of 25-ANE. Competitive binding assays of 25-OH-D3 and 25-ANE with rat serum demonstrated that 25-ANE competes for the 25-OH-D3 binding site in rat serum vitamin D binding protein (rDBP). On the other hand, UV exposure of a sample of purified rat DBP (rDBP), preincubated in the dark with 3H-25-ANE, covalently labeled the protein. However, very little covalent labeling was observed in the absence of UV light or in the presence of a large excess of 25-OH-D3. These results provide strong evidence for the covalent labeling of the 25-OH-D3 binding site in rDBP by 3H-25-ANE.

Affinity Labels↗
PubMed1991

Vitamin D analogs with low affinity for the vitamin D binding protein: enhanced in vitro and decreased in vivo activity.

The affinity of 1 alpha,25-dihydroxyvitamin D3 [1 alpha,25-(OH)2D3] and analogs with side-chain modifications [MC 903 or calcipotriol, MC 1147 or 24,24-dihomo-1 alpha,25-(OH)2D3 and 1,25-(OH)2-16ene-23yne-D3] for the vitamin D receptor and the serum vitamin D binding protein (DBP) were compared. The affinity of MC 903 for the receptor from chick and rat duodenum or from human peripheral blood mononuclear cells or HL-60 cells varied between 60 and 100% relative to the affinity of 1,25-(OH)2D3. The relative affinity of 1,25-(OH)2-16ene-23yne-D3 and MC 1147 varied for the same receptors between 45-70 and 3.5-25%, respectively. The relative affinity of MC 903 for human DBP was 30-fold decreased, whereas the two other analogs did not bind to DBP at all even in more than 1000-fold excess. The in vitro biologic activity of 1 alpha,25-(OH)2D3 on phytohemagglutinin-stimulated normal human lymphocyte proliferation was markedly inhibited by the addition of physiologic amounts of DBP to the cell culture medium. No such inhibition was observed when MC 903 or 1147 was evaluated similarly. DBP therefore reversed the rank order of the in vitro potency of these analogs. Intramuscular injections for 10 consecutive days to vitamin D-deficient chicks demonstrated a greater than or equal to 100-fold lower biologic activity of MC 903, MC 1147, and 1,25-(OH)2-16ene-23yne-D3 compared to that of 1 alpha,25-(OH)2D3 as evaluated by serum calcium and osteocalcin concentrations, as well as by duodenal calbindin D28K and bone calcium content.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Short-term course of 1,25(OH)2D3 stimulates osteoblasts but not osteoclasts in osteoporosis and osteoarthritis.

We investigated the effect of short-term, 1,25-dihydroxyvitamin D3 therapy (4 micrograms/day for 4 days) on calcium metabolism in 27 postmenopausal women (11 cases with osteoporosis and 16 cases with osteoarthritis). Bone mass at the axial and appendicular skeleton was higher in osteoarthritis than in osteoporosis. Initial values of calcium metabolism were similar. Osteoporotic and osteoarthritic patients responded with a similar significant increase in serum osteocalcin (+61% and +54%, respectively), fasting urinary calcium excretion (+178% and +124%, respectively) and 24 hour calcium excretion (+148% and +142%, respectively). Parathyroid hormone (PTH) levels decreased significantly in both groups (-30% and -18%, respectively). Osteoclastic bone resorption, evaluated by urinary hydroxyproline excretion, was not stimulated in either group. We conclude that in osteoporosis and also in osteoarthritis (1) 1,25-dihydroxy-vitamin D3 (1,25(OH)2D3) stimulation of osteoblast function is similar in production of osteocalcin; (2) the vitamin D target tissues react adequately to 1,25(OH)2D3 stimulation; (3) short-term high dose of 1,25(OH)2D3 does not stimulate bone resorption; and (4) the differences in bone mass between osteoarthritis and osteoporosis are not related to an alteration of the responsiveness to stimulation by 1,25 (OH)2D3.

Aged↗
PubMed1991

Seasonal variation in bone metabolism in young healthy subjects.

Serum vitamin D metabolites and urinary calcium excretion; parameters of bone formation (serum alkaline phosphatase, serum osteocalcin); parameters of bone resorption (24 hour hydroxyprolinuria, 2 hour fasting urinary hydroxyproline/creatinine ratio); and parameters of cortical and trabecular bone density, parathyroid hormone (iPTH, COOH terminal assay), and serum minerals (calcium, phosphorus) were followed serially in 55 young adults (21 women and 34 men) from December 1985 until January 1987 at four different times during the year. The effect of a low-dose cyclooxygenase inhibitor (piroxicam 5 mg daily) on the same parameters of bone density and bone turnover when given from December until May, was also evaluated in this study. At the end of the treatment period parameters of bone turnover and bone density were comparable between placebo and piroxicam-treated groups. Therefore, the results of all subjects were pooled in order to investigate seasonal variation. In both sexes, seasonal variation was found not only for 250HD3 but also for 1,25(OH)2D3, serum calcium and phosphorus, urinary calcium excretion, and for bone density at the lumbar spine. Parameters of bone formation (serum osteocalcin and alkaline phosphatase), bone resorption (24 hour urinary hydroxyprolinuria and fasting urinary hydroxyproline/creatinine ratio) and PTH were influenced by this seasonal variation. We conclude that in young adults, a significant seasonal variation occurs, with low winter and high summer values, for serum 25 and 1,25(OH)2D3 for urinary calcium apparently without important influence on parameters of bone turnover or parathyroid activity and for lumbar spine density. Treatment with a low-dose cyclooxygenase inhibitor was without influence on the observed changes.

Adult↗
PubMed1991

Distribution of group-specific component/vitamin-D-binding protein subtypes in Saudi Arabia.

The distribution of group-specific component (Gc) phenotypes and the Gc allele frequencies were investigated in 1,082 individuals from five different provinces of Saudi Arabia by the combined use of isoelectric focusing and immunofixation. Between provinces variations in the Gc allele frequencies were found. Gc1S decreased and Gc1F increased from the northwest to the southeast in Saudi Arabia. The overall frequencies in Saudi Arabia were 0.236 for Gc1F, 0.610 for Gc1S, 0.150 for Gc2 and 0.004 for rare alleles. The observed frequencies did not differ significantly from those found in other population samples from the Middle East. In nine samples rare Gc variants were found.

Alleles↗
PubMed1991

Growth hormone and bone.

Bone and mineral metabolism is regulated by numerous hormones and local growth factors. Growth hormone (GH), directly and/or indirectly through local IGF1 and IGF2 production, stimulates bone turnover as it increases osteoblast number and function. It also stimulates bone resorption through unknown mechanisms. GH (by IGF1) stimulates the renal 25-hydroxyvitamin D-1 alpha-hydroxylase activity and therefore enhances calcium and phosphate absorption in the intestine. It also increases the maximal renal tubular reabsorption of phosphate. The overall effect of GH on bone therefore seems beneficial as bone mass is increased together with the lean body mass. Large excess of GH, however, also increases the risk of osteoarthritis and has other serious metabolic side effects, so that its place in the treatment/prevention of osteoporosis is yet to be defined.

Animals↗
PubMed1991

Attenuated rise of 1,25 (OH)2 vitamin D3 in response to parathyroid hormone in patients with incipient renal failure.

Despite elevated parathyroid hormone (PTH) levels, low normal or diminished serum 1,25(OH)2D3 concentrations are found in patients with incipient renal failure. To further assess (indirectly) the reserve capacity of renal production of 1,25(OH)2D3 we studied 9 patients with incipient or moderate renal failure (inulin clearance 31-68ml/min/1.73 m2) and 9 controls, using a novel stimulation test. We measured 1,25 (OH)2D3 levels, free 1,25(OH)2D3 index, cAMP excretion, calciuria and phosphaturia before and after infusion of 2 x 400 U of human (h) PTH (1-38). Baseline 1,25(OH)2D3 levels were not significantly different in patients (42.5 pg/ml, 21.6-51.1) compared with controls (45.0 pg/ml, 37.4-67.3). After infusion of hPTH(1-38), however, median increase in 1,25(OH)2D3 was only +25% versus +86% in controls, despite a greater proportional increase in cAMP/GF ratio. The data suggest subnormal stimulation of renal 1,25(OH)2D3 production in response to exogenous PTH in most patients with incipient renal failure. This may reflect partial exhaustion of biosynthetic reserve capacity.

Adult↗
PubMed1991

Comparison of intact, midregion, and carboxy terminal assays of parathyroid hormone for the diagnosis of bone disease in hemodialyzed patients.

The predictive value of three different RIAs of PTH for the diagnosis of the histological type of bone disease has been compared in 24 asymptomatic patients on chronic hemodialysis who had never been exposed to aluminum intoxication and who agreed to have a bone biopsy after double tetracycline labeling. The serum concentrations of PTH were measured using a two-site immunoradiometric assay for intact PTH(1-84) and region specific assays directed against the C-terminal (53-84) fragment or the midregion (44-68) of the molecule. The bone histomorphometric analysis showed that six patients had nonaluminic adynamic bone disease with low bone formation rate (BFR), eight had mild hyperparathyroidism characterized by increased bone resorption and normal BFR, nine had severe hyperparathyroidism with increased BFR, and only one had true osteomalacia with increased osteoid seam thickness. All PTH assays correlated with the various parameters of bone resorption and bone formation and were able to differentiate the histological type of bone disease only when groups of patients were considered. For classifying individual patients into severe hyperparathyroidism and adynamic bone disease groups, the intact PTH assay had the best predictive value with a sensitivity of 100% and a specificity of at least 70%. A nonaluminic adynamic bone disease was observed in more than 50% of the patients who had normal intact PTH levels (6/11). It is concluded that the intact PTH measurement is superior to C-terminal and midregion assays for the prediction of the histological type of bone disease in hemodialyzed patients and should be of considerable value to adapt their treatment in order to avoid the emergence of both severe hyperparathyroidism and adynamic bone disease. In the absence of aluminum intoxication it seems that maintaining intact PTH concentrations 1 to 1.5 times the upper limit of normal would correspond to the best bone histology.

Bone Diseases↗
PubMed1990

Improvement of gastric emptying in diabetic gastroparesis by erythromycin. Preliminary studies.

Erythromycin mimics the effect of the gastrointestinal polypeptide motilin on gastrointestinal motility, probably by binding to motilin receptors and acting as a motilin agonist. Erythromycin may thus have clinical application in patients with disturbances of gastroduodenal motility, such as diabetic gastroparesis. To examine this possibility, we studied the effect of erythromycin on gastric emptying in 10 patients with insulin-dependent diabetes mellitus and gastroparesis. We studied the emptying of liquids and solids simultaneously on separate days after the intravenous administration of erythromycin (200 mg) or placebo, using a double-isotope technique and a double-blind, crossover design. Erythromycin shortened the prolonged gastric-emptying times for both liquids and solids to normal. For example, 120 minutes after the ingestion of a solid meal, mean (+/- SE) retention was 63 +/- 9 percent with placebo and 4 +/- 1 percent with erythromycin, as compared with 9 +/- 3 percent in 10 healthy subjects. The corresponding values 120 minutes after the ingestion of a liquid meal were 32 +/- 4, 9 +/- 3, and 4 +/- 1 percent, respectively. Gastric emptying also improved, but to a lesser degree, in the 10 patients after four weeks of treatment with oral erythromycin (250 mg three times a day). These preliminary results suggest that erythromycin may have therapeutic value in patients with severe diabetic gastroparesis.

Adult↗
PubMed1990

Effects of vitamin D-binding protein on bone resorption stimulated by 1,25 dihydroxyvitamin D3.

Vitamin D and its metabolites are tightly bound to the serum vitamin D-binding protein (DBP) and only the free hormone is considered to be physiologically active. On the other hand, DBP could interact with cell membranes and even favor its intracellular entry. The present study was undertaken to examine the effects of DBP on bone resorption stimulated by 1,25-dihydroxyvitamin D3 [1,25(OH)2D3]. Forelimb bones from 19-day-old fetal rats were cultured for 5 days in the presence of purified human or rat serum albumin (hSAP or rSAP) and 1,25(OH)2D3, with or without human or rat DBP (hDBP or rDBP). Bone resorption was assessed by measuring the release of previously incorporated 45Ca. We found that the resorptive response to 1,25(OH)2D3 was minimally altered by hDBP (5 microM). The minimal effects of hDBP on 1,25(OH)2D3 activity on rat bones might be explained by a 6-fold lower affinity of hDBP (1.1 x 10(7) M-1) than rDBP (5.9 x 10(7) M-1) for 1,25(OH)2D3 or by species differences in cellular recognition of DBP. In a homologous rat system, however, rDBP at low (0.5 microM) or physiological (5 microM) concentration significantly decreased 1,25(OH)2D3-induced bone resorption. These data therefore support the hypothesis that free rather than DBP-bound 1,25(OH)2D3 is physiologically important.

Animals↗
PubMed1990

Particles from dialysis tubing stimulate interleukin-1 secretion by macrophages.

Loading of tissue macrophages with dialysis-tubing-derived particles may occur during chronic haemodialysis. Previous studies have demonstrated that these particle-laden macrophages release significant quantities of prostaglandins. In these experiments, the effects of dialysis-tubing-particle loading on the release of the central inflammatory mediator, interleukin 1 (IL 1), was examined. Rats received daily injections of silicone or polyvinylchloride (PVC) particles, and were compared to animals given saline alone. The silicone and PVC groups received a total of 3 x 10(9) particles over a 4-week period. Non-stimulated peritoneal macrophages from control animals released a median of 4.1 (range 1.2-10.3) U IL 1 per 10(6) cells. In contrast, macrophages from silicone- and PVC-loaded animals spontaneously released high levels of IL 1 (median 21.8; range 10-36.7) and 94 (range 36-336) U per 10(6) cells respectively). Following in vitro stimulation with bacterial lipopolysaccharide (LPS), peritoneal macrophages from silicone- and PVC-treated animals released large amounts of IL 1 (median 538 (range 359-2017) U and median 653 (range 326-1134) U per 10(6) cells, respectively) as compared to LPS-stimulated macrophages from control animals (median 332 (range 130-306) U per 10(6) cells]. Zymosan or LPS stimulation of splenic cells from silicone- and PVC-loaded animals also secreted increased quantities of IL 1 as compared to controls. The chronic loading of tissue macrophages in dialysis patients with tubing-derived particles may result in augmented release of IL 1, with subsequent activation of inflammatory processes.

Animals↗
PubMed1990

Osteoporosis and diabetes: lessons from the diabetic BB rat.

The spontaneously diabetic BB rat was used to study the effect of diabetes on bone. Short-term diabetes (3-4 weeks) resulted in a state of low bone turnover, characterized by a severe decrease in osteoblast/osteoid surface and bone mineral apposition rate on histology, and in serum osteocalcin concentrations. If diabetes was long-term (12 weeks), the parameters of low bone formation were associated with histological evidence of osteoporosis and a decreased bone strength; the relative bone calcium concentration remained normal in diabetes. We conclude that long-standing diabetes results in a low-turnover osteoporosis.

Animals↗
PubMed1990

Changes in biochemical markers of osteoblastic activity during the menstrual cycle.

Serum levels of osteocalcin [OC; bone Gla protein (BGP)] and bone alkaline phosphatase (B-AP) are both correlated to osteoblastic activity, which may be regulated by several hormones, including estrogen, 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3], and PTH. Estrogen shows reproducible variations during the menstrual cycle, while available data on variations in serum 1,25-(OH)2D3 and serum immunoreactive PTH show midcyclic increases or no changes. In the present study we evaluated osteoblastic activity by measuring serum OC and B-AP during the menstrual cycle in eight healthy women, aged 20-47 yr. The cycles were synchronized by LH peaks, and follicular and luteal periods were normalized by lengths. Repeated measures analysis of variance showed that serum OC varied significantly (P less than 0.05), with highest levels during the luteal period. Although the same pattern was seen for serum B-AP, the variation just failed to reach significance (P less than 0.10), but the mean level was significantly higher during the luteal than during the follicular period (P less than 0.05). Gonadotropins and ovarian sex hormones showed significant variations. There were no significant changes in serum vitamin D-binding protein, serum total and free 1,25-(OH)2D3 index, or serum immunoreactive PTH-(1-84), but serum levels of somatomedin-C showed a significant variation, with the highest level during the luteal period (P less than 0.05). Blood levels and urinary excretion of minerals exhibited no significant variations. Cross-correlation studies between OC and estradiol showed the highest correlation coefficient, when OC was lagged about 7 days after estradiol (r = 0.69; P less than 0.05). Moreover, a high correlation was found between OC and somatomedin-C when matched at concurrent time points (r = 0.76; P less than 0.01). No significant correlations were found between the other calcium-regulating hormones and OC when matched at concurrent time points. In conclusion, we found a significant effect of the menstrual cycle on the serum levels of two osteoblastic bone markers, OC and B-AP. The changes indicated that osteoblastic activity is higher during the luteal period. However, whether the changes are caused by direct or indirect effects of the fluctuations in calciotropic hormones is still unresolved.

Adult↗
PubMed1990

Bone and mineral metabolism in BB rats with long-term diabetes. Decreased bone turnover and osteoporosis.

The effect of long-term diabetes mellitus on bone and mineral metabolism was studied in BB rats. Diabetic rats were treated with 1 U of long-acting insulin every other day for 12 wk and compared with nondiabetic littermates. Urinary calcium excretion was increased greater than 10-fold, but serum total and diffusible calcium remained normal. Serum concentrations of both 1 alpha, 25-dihydroxyvitamin D3 and vitamin D-binding protein were significantly decreased in diabetic rats. The intestinal calbindin-D 9K concentration was decreased by nearly 50%, and active duodenal calcium absorption was totally abolished. Trabecular bone volume measured in the tibial metaphysis was decreased by 44%, and the osteoblast and osteoid surfaces were less than 10% of values observed in control rats, whereas the osteoclast surface was unchanged by diabetes. The daily bone formation (bone mineral apposition rate) measured by labeling twice with calcein was decreased by 86% in diabetic rats. The serum concentration of osteocalcin, a biochemical marker of osteoblast function, was similarly decreased (mean +/- SE 23 +/- 3 and 62 +/- 4 micrograms/L in diabetic [n = 15] and nondiabetic [n = 15] rats, respectively). Serum osteocalcin was significantly correlated with the serum concentration of insulinlike growth factor I (r = 0.89, P less than 0.001). Bone strength measured as the energy needed to fracture the femur was markedly decreased (5.3 +/- 1.4 and 8.4 +/- 1.3 N.m.degree in diabetic and nondiabetic rats, respectively; P less than 0.01). These histological, chemical, and biomechanical data clearly indicate that long-standing diabetes in BB rats results in severe low-turnover osteoporosis probably related to decreased osteoblast recruitment and/or function.

Animals↗
PubMed1990

Immunoradiometric assay of parathyrin with polyclonal and monoclonal region-specific antibodies.

In this immunoradiometric assay (IRMA) of parathyrin (PTH) a polyclonal anti-amino-PTH(1-34) is the capture antibody and a radiolabeled monoclonal anti-hPTH(44-68) is the second antibody. Gel filtration of serum from a hyperparathyroid patient yielded only a single peak of PTH, corresponding to the elution position of synthetic PTH(1-84). Healthy elderly individuals (ages 78 +/- 5 y, mean +/- SD, n = 45) had PTH concentrations (21 +/- 13 ng/L) not significantly higher than those from healthy younger (38 +/- 11 y) adults (20 +/- 8 ng/L, n = 94). Assay results agreed well with those obtained with a carboxyl-terminal PTH assay both in normal subjects (r = 0.63, P less than 0.001) and in patients with primary hyperparathyroidism (r = 0.59, P less than 0.001). Both assays equally discriminated patients with surgically confirmed primary hyperparathyroidism from normal individuals, but the PTH(1-84) IRMA also allowed a nearly absolute discrimination between normal subjects and patients with primary hypoparathyroidism (undetectable serum PTH in 18 of 21 cases) and secondary hypoparathyroidism (caused by hypercalcemia that was caused by a malignant tumor, PTH 1.3 +/- 1.3 ng/L, n = 32). Moreover, the PTH(1-84) IRMA is more sensitive (detection limit in serum, 0.8 ng/L) and easier and quicker to perform than the carboxyl-terminal assay.

Adult↗
PubMed1990

Osteocalcin is vitamin D-dependent during the perinatal period in the rat.

The vitamin D-dependence of renal calbindin D-28K and osteocalcin during the perinatal period was studied in fetuses (days 18 and 21) and neonates (days 2, 12, 17 and 22) of rats fed either a standard diet (0.85% Ca-0.7% P; "high Ca-P diet" rats) or a mildly Ca-P restricted diet (0.2% Ca-0.2% P; "low Ca-P diet" rats). Body weight and plasma calcium levels were identical in both groups. Plasma 1,25(OH)2D concentrations were markedly higher in the low Ca-P diet rats at all stages of fetal and neonatal life (in 22-day-old neonates: 536 +/- 58 pg/ml versus 126 +/- 12 pg/ml). 1,25(OH)2D concentrations increased between day 18 and 21 of fetal life, remained constant between day 21 of fetal and day 12 of neonatal life, and increased sharply between day 12 and 17 in both groups; after day 17, 1,25(OH)2D concentrations increased further in pups fed the low Ca-P diet. Renal calbindin D-28K reached peak concentrations on day 12 of neonatal life; calbindin D-28K levels were similar in the high and low Ca-P diet rats at all stages of perinatal development. Plasma osteocalcin levels increased steadily during the perinatal period; at most stages of perinatal life, and already from the fetal period was osteocalcin higher in the low Ca-P diet rats than in the high Ca-P diet rats (in 22-day-old pups: 1106 +/- 47 ng/ml versus 429 +/- 14 ng/ml). Femoral osteocalcin concentrations were also increased in fetal and early neonatal (days 2 and 12) low Ca-P diet rats, while the femoral calcium content and concentration of these rats were decreased in the late neonatal period (days 12, 17 and 22). These studies indicate that osteocalcin is vitamin D-dependent in the fetal and neonatal rat.

Animals↗